Conjugative mapping of pyruvate, 2-ketoglutarate, and branched-chain keto acid dehydrogenase genes in Pseudomonas putida mutants.
Conjugative mapping of pyruvate, 2-ketoglutarate, and branched-chain keto acid dehydrogenase genes in Pseudomonas putida mutants.
复制标题
恶臭假单胞菌突变体中丙酮酸、2-酮戊二酸和支链酮酸脱氢酶基因的共轭作图。
DOI：
10.1128/jb.162.1.203-208.1985
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发表时间：
1985
影响因子：
3.2
通讯作者：
Sokatch,JR
中科院分区：
文献类型：
--
作者：
Sykes,PJ;Menard,J;McCully,V;Sokatch,JR
支链酮酸脱氢酶是恶臭假单胞菌支链氨基酸分解代谢共同途径中的一种酶，是催化支链酮酸氧化脱羧基的多酶复合体。本研究的目的是分离具有该酶和其他酮酸脱氢酶突变的菌株，并定位恶臭假单胞菌染色体上的突变位置。分离到两株丙酮酸和两株2-酮戊二酸脱氢酶基因突变菌株，并通过生化分析鉴定了缺陷亚基。通过使用重组XYL-K质粒来介导接合，这些突变被定位到一系列营养缺乏症和其他分解代谢突变。记录的最后一次进入时间约为35分钟，数据与单一进入点一致。影响E1、E1加E2和E3亚基的支链酮酸脱氢酶突变大约在35分钟被定位。在共同途径中受影响的另一株菌株缺乏支链氨基酸转氨酶，突变被定位在16分钟。两个丙酮酸脱氢酶突变体，一个缺乏E1，另一个缺乏E1+E2，在22分钟绘制出突变图谱。影响E1亚基的2-酮戊二酸脱氢酶突变被定位在12分钟。分离到了一个E3缺失的2-酮戊二酸脱氢酶突变体，但该突变被证明过于泄漏而无法定位。
Branched-chain keto acid dehydrogenase, an enzyme in the common pathway of branched-chain amino acid catabolism of Pseudomonas putida, is a multienzyme complex which catalyzes the oxidative decarboxylation of branched-chain keto acids. The objective of the present study was to isolate strains with mutations of this and other keto acid dehydrogenases and to map the location of the mutations on the chromosome of P. putida. Several strains with mutations of branched-chain keto acid dehydrogenase, two pyruvate and two 2-ketoglutarate dehydrogenase, were isolated, and the defective subunits were identified by biochemical analysis. By using a recombinant XYL-K plasmid to mediate conjugation, these mutations were mapped in relation to a series of auxotrophic and other catabolic mutations. The last time of entry recorded was at approximately 35 min, and the data were consistent with a single point of entry. Branched-chain keto acid dehydrogenase mutations affecting E1, E1 plus E2, and E3 subunits mapped at approximately 35 min. One other strain affected in the common pathway was deficient in branched-chain amino acid transaminase, and the mutation was mapped at 16 min. The mutations in the two pyruvate dehydrogenase mutants, one deficient in E1 and the other deficient in E1 plus E2, mapped at 22 minutes. The 2-ketoglutarate dehydrogenase mutation affecting the E1 subunit mapped at 12 minutes. A 2-ketoglutarate dehydrogenase mutant deficient in E3 was isolated, but the mutation proved too leaky to map.